The overall goal of this program is to develop an understanding of the role(s) played by sialoglycoproteins in mammary tumors. This project will concentrate on a large, predominantly mucin-type sialoglycoprotein (ASGP-1) found in ascites forms of the 13762 mammary adenocarcinoma, but not in solid or cultured forms of the tumor. ASGP-1 molecules found in xenotransplantable (MAT-C1) and species specific (MAT-B1) forms of the tumor have been purified to homogeneity and found to have essentially identical amino acid ccmpositions, but very different carbohydrate compositions. MAT-B1 ASGP-1 can be labeled with sulfate, but MAT-C1 ASGP-1 shows little labeling. Since differences in sulfation and sialic acid content suggest differences in the control of the biosynthesis of ASGP-1 oligosaccharides, we will investigate structures of the oligosaccharides released from ASGP-1 by alkaline borohydride. We will also investigate the nature of the polypeptide chain by classical degradation techniques. The mode of association of ASGP-1 with membranes will be studied by detergent extraction and reconstitution with lipids. Antibodies against ASGP-1 and its nonglycosylated polypeptide portion will be prepared. Using cell fractionation techniques, metabolic labeling and the antibodies, we will investigate "shedding" of ASGP-1 by the tumor cells in vitro and in vivo. Rates of component release and the nature of released components will be determined by labeling, centrifugation, electrophoresis and gel filtration. Turnover of ASGP-1 will be analyzed by pulse-chase double label experiments and compared to turnover of other cellular constituents. The presence of ASGP-1 in solid and cultured tmors will be assessed by labeling and immunological analyses. The ability of clones of the cultured tumor to produce ASGP-1 will be tested. These studies should provide considerable insight into the molecular mechanisms of expression of ASGP-1 that may be important to a role in allowing the tumors to evade immune surveillance.